Nonnucleoside Inhibitor of Measles Virus RNA-Dependent RNA Polymerase Complex Activity

Abstract

Paramyxoviruses comprise several major human pathogens. Although a live-attenuated vaccine protects against measles virus (MV), a member of the paramyxovirus family, the virus remains a principal cause of worldwide mortality and accounts for approximately 21 million cases and 300,000 to 400,000 deaths annually. The development of novel antivirals that allow improved case management of severe measles and silence viral outbreaks is thus highly desirable. We have previously described the development of novel MV fusion inhibitors. The potential for preexisting or emerging resistance in the field constitutes the rationale for the identification of additional MV inhibitors with a diverse target spectrum. Here, we report the development and implementation of a cell-based assay for high-throughput screening of MV antivirals, which has yielded several hit candidates. Following confirmation by secondary assays and chemical synthesis, the most potent hit was found to act as a target-specific inhibitor of MV replication with desirable drug-like properties. The compound proved highly active against multiple primary isolates of diverse MV genotypes currently circulating worldwide, showing active concentrations of 35 to 145 nM. Significantly, it does not interfere with viral entry and lacks cross-resistance with the MV fusion inhibitor class. Mechanistic characterization on a subinfection level revealed that the compound represents a first-in-class nonnucleoside inhibitor of MV RNA-dependent RNA polymerase complex activity. Singly or in combination with the fusion inhibitors, this novel compound class has high developmental potential as a potent therapeutic against MV and will likely further the mechanistic characterization of the viral polymerase complex.

Independent Structural Domains in Paramyxovirus Polymerase Protein

“…Novel nucleoside and allosteric inhibitors of hepatitis C virus polymerase, for instance, are at different stages of clinical evaluation and show strong drug potential (83)(84)(85). Experimentally tested allosteric blockers of Paramyxovirus polymerase likewise combine high potency with minimal cytotoxicity, opening a desirably large therapeutic window (86,87). High resolution information about the Mononegavirales polymerase core structure would pave the path for structure-guided de novo drug design efforts, the informed optimization of existing inhibitor scaffolds, and the proactive design of inhibitor protected against rapid viral escape from inhibition.…”

Respiratory Syncytial Virus Attachment Glycoprotein Contribution to Infection Depends on the Specific Fusion Protein

“…To further explore the functional differences between 2-20 G and A2 G, we tested whether virus entry into host cells would be differentially affected by the removal of A2 or 2-20 G protein. We quantified entry kinetics of all four viruses in BEAS-2B cells by the use of a citric acid wash entry assay (25,28). kRSV-A2GA2F had the fastest entry kinetics, followed by kRSV-GstopA2F (Fig.…”

“…This assay was performed as described previously with some modifications (25). BEAS-2B cells (70% confluent in 12-well plates) were placed on ice for 5 min and washed once with ice-cold PBS before addition of virus at an MOI of 1.0.…”

Target Analysis of the Experimental Measles Therapeutic AS-136A

“…Assessment of an ϳ34,000-entry small-molecule diversity set has yielded a novel compound class with target-specific nanomolar antiviral activity (57). Hit-to-lead chemistry has further advanced this compound class (53,54), rendering it one of the most promising experimental MeV drugs identified.…”